1. Technical Field
The present invention relates to a reagent for a high sensitive immunoassay method, an immunoassay method using said reagent and a reagent kit comprising said reagent.
2. Related Art
It has been known that some biological substances exist both in a free form and a bound form. A representative example thereof is the prostate-specific antigen (PSA) which has been known to exist in a bound form of PSA and α1-antichymotrypsin (ACT) in blood, as well as in a free form of PSA itself. Since such substances exhibit a change in a ratio of the free form and the bound form depending on an amount of substance bound with the free form as well as in vivo circumstances, both forms should be assayed in the assay of such substances. However, there is a problem that in an immunoassay of such substances by applying antigen-antibody reaction, a high precise assay is difficult due to different binding rate of the antibody to the free form and the bound form. In order to solve such the problem, JP-A-09-234068 proposes an assay reagent for equimolar reaction with both free form and bound form, and an assay method using the same are proposed. Since this method is, however, the assay method by ELISA, it can be possible to assay only in the specific equipment and is impossible to apply in the common autoanalyzer. Although the latex turbidimetry is known as a high sensitive immunoassay method by applying the common autoanalyzer, there is also a problem that the equimolar assay of both free form and bound form is difficult in such a method, even if the latex sensitized (immobilized) with the antibody which exhibits equimolar reaction in ELISA is used. Owing to these problems, a development of a method is demanded, which can assay the substance existing in the free form and the bound form in the organism and can be applied in the common autoanalyzer.
In JP-A-10-123137, an immunoassay method is disclosed, in which at least two types of different monoclonal antibodies for a specific antigen are carried by an insoluble carrier such as latex and the insoluble carrier having different mean particle diameter is used. However, in this method, the free form and the bound form in the specimen are not assayed.